Abstract
Nuclear egress is an essential process in the replication of human cytomegalovirus (HCMV), as it enables the migration of newly formed viral capsids from the nucleus into the cytoplasm. Inhibition of the HCMV core nuclear egress complex (core NEC), composed of viral proteins pUL50 and pUL53, has been proposed as a potential new target for the treatment of HCMV infection and disease. Here, we present a new type of small molecule inhibitors of HCMV core NEC formation, which inhibit the pUL50-pUL53 interaction at nanomolar concentrations. These inhibitors, i.e., verteporfin and merbromin, were identified through the screening of the Prestwick Chemical Library® of approved drug compounds. The inhibitory effect of merbromin is both compound- and target-specific, as no inhibition was seen for other mercury-organic compounds. Furthermore, merbromin does not inhibit an unrelated protein–protein interaction either. More importantly, merbromin was found to inhibit HCMV infection of cells in three different assays, as well as to disrupt HCMV NEC nuclear rim formation. Thus, while not being an ideal drug candidate by itself, merbromin may serve as a blueprint for small molecules with high HCMV core NEC inhibitory potential, as candidates for novel anti-herpesviral drugs.
Highlights
Human cytomegalovirus (HCMV) is a ubiquitous human β-herpesvirus with a seroprevalence ranging from 40% to 95% in various regions of the world, establishing a life-long latent infection
The viral genome equivalents of the supernatants of HCMV-infected Human foreskin fibroblast (HFF) treated with merbromin at concentrations ranging from 0.3 to 10 μM were quantified by Quantitative Polymerase Chain Reaction (qPCR)
Cytotoxicity and Selectivity of Merbromin As both verteporfin and merbromin have been associated with a considerable degree of toxicity, we addressed the in vitro cytotoxicity of both compounds, prior to testing them in cell-based HCMV infection assays
Summary
Human cytomegalovirus (HCMV) is a ubiquitous human β-herpesvirus with a seroprevalence ranging from 40% to 95% in various regions of the world, establishing a life-long latent infection. A similar N-terminal pUL53 peptide was previously shown to interfere, albeit at 100-fold higher concentrations, with the pUL50-pUL53 interaction [15] Based on these data, we have proposed inhibition of the pUL50-pUL53 interaction as a novel antiviral strategy against HCMV infections. Due to its size (molecular weight: approximately 4 kDa), as well as its susceptibility to proteolytic degradation, the pUL53 hook peptide is a less than ideal candidate for inhibition of the pUL50-pUL53 interaction in vivo This becomes even more apparent considering that a successful inhibitor of this interaction has to be taken up by the cell, as well as to penetrate the nuclear membrane. In which “100%” is a sample without inhibitor, “blank1” is a sample without pUL53, and “blank2” is a sample without pUL53 and without inhibitor
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